Project Summary Mother-to-child transmission (MTCT) of HIV remains a crisis in resource-limited countries where HIV is prevalent. Approximately 200,000 MTCTs of HIV occur annually, with as many as half of infections being due to exposure via breastmilk (BM) [1, 2]. Yet, only ~10-15% of infants breastfed by HIV-infected mothers actually become infected, suggesting a strong protective effect of BM itself [1-5]. Though multiple studies have demonstrated the protective effect of human BM against HIV MTCT [1-10], the contribution of the milk?s cellular component has been relatively overlooked, despite evidence that maternal leukocytes are functional beyond the sites of ingestion [11-15]. The only clinical HIV vaccine trial to show efficacy, RV144, and many other studies have correlated activities mediated by the constant (Fc) Ab domain with protection from HIV acquisition and this is documented similarly with other pathogens [16-29]. Though demonstrated as necessary for the clearance of numerous viral infections, one essential Fc-mediated response--Ab-dependent cellular phagocytosis (ADCP)--has been relatively understudied in the context of HIV, particularly in the case of prevention of MTCT [30-37]. Colostral phagocytes can perform ADCP of bacteria, parasites and yeast opsonized with maternal Abs; however, this has not been studied with regard to HIV or HIV-infected cells [38- 44]. Furthermore, the potential contribution of ADCP to protection from MTCT of HIV has not been studied with regard to impact of phagocytic target size (e.g., cell-free and cell-associated virus), or the effects of the dynamic leukocyte composition of BM over the lactation period [45-50]. Only conflicting and/or small studies have been conducted regarding the relevance of Ab subclass in Fc-mediated activity, especially in BM [16, 42, 51-57]. Given the substantial gap in present knowledge of the potential contribution of ADCP activity by BM phagocytes to prevention of MTCT of HIV, it is critical to develop a multidimensional, comprehensive understanding of ADCP by the relevant primary cells in BM. The proposed study aims to fill this knowledge gap. AIM 1 will address the impact of phagocytic target size/type on ADCP by BM cells, AIM 2 will address the impact of Ab class on ADCP by BM cells, and AIM 3 will address the impact of BM maturation over time on ADCP activity. These data will allow the field to better understand the potential contribution of ADCP mediated by BM cells to the reduction of MTCT of HIV, and may well be applicable to other pathogens that threaten infants over the course of lactation.